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Miscreant forms of tau are known to start trouble when they wander away from axons and into presynaptic terminals. In the February 1 Neuron, researchers report that the synaptic vesicle protein synaptogyrin-3 engages tau during this walkabout. Researchers led by Patrik Verstreken at KU Leuven in Belgium used both rodent and insect models to discover that, in the terminal space, tau tethers itself to synaptic vesicles via synaptogyrin-3, then cross-links the neurotransmitter packages, hindering their release. The researchers proposed that this presynaptic dysfunction could happen early in AD, and that tau’s association with synaptic vesicles might even facilitate the transfer of tau between neurons.

“This suggests a very interesting mechanism of tau-mediated neurotoxicity,” commented Markus Zweckstetter of the Max Planck Institute for Biophysical Chemistry in Munich. Zweckstetter added that as a link to tau’s association with synaptic vesicles, synaptogyrin-3 could be an important player in tau-mediated neuronal damage.

Owing to its association with microtubules, healthy tau predominantly resides in axons. However, in pathological situations such as AD, researchers have spotted tau accumulating in the somatodendritic compartment, which has prompted abundant research on the evils of dendritic tau (e.g. Sep 2010 news; Jan 2011 news; Jul 2017 news).

However, recent studies have indicated that tau can wreak havoc closer to their natural habitat, that is, within presynaptic terminals that line axons. In 2015, researchers reported that tau accumulation in presynapses caused dysfunction and loss of synapses (Apr 2015 news). More recently, researchers in Verstreken’s lab closed in on an explanation for this. They reported that tau latched onto synaptic vesicles via its N-terminus and to actin filaments with its C-terminus, effectively cross-linking the vesicles and hindering their release during signaling (Apr 2017 conference news; Zhou et al., 2017).

What links tau to synaptic vesicles in the first place? First author Joseph McInnes, now at Baylor College of Medicine in Houston, Texas, and colleagues addressed this question in the current study. Before dealing with the nitty-gritty of tau’s relationship with vesicles, they wanted to confirm that tau actually associates with vesicles in AD patient brains. Their previous study had only shown hyperphosphorylated tau in the presynapse of human postmortem samples, but did not nail down its association with vesicles there. They fractionated postmortem brain tissue from 15 AD patients and 13 non-demented controls. In the synaptic vesicle fraction, they found two- to threefold more tau in AD samples than controls, and also that this vesicle-associated tau was hyperphosphorylated and oligomeric. This suggested the association of tau with synaptic vesicles was pathologically relevant.

The researchers next used monomeric, soluble human tau as bait to fish for interaction partners on synaptic vesicles isolated from mouse brains. By a process of molecular elimination they narrowed down the search to proteins, then to transmembrane proteins. Using proteomics, they determined that synaptic vesicle transmembrane proteins bound to full-length, but not N-terminally truncated tau, and zeroed in on synaptogyrin-3. The protein has four transmembrane domains with both its N- and C-termini exposed to the cytoplasm. Its function is unknown.

To investigate how synaptogyrin-3 affects tau localization in neurons, the researchers turned to hippocampal neurons from PS19 mice, which express the P301S mutant form of tau that causes frontotemporal dementia. As reported before, P301S tau co-localized with presynaptic terminal markers in punctate clusters. When the researchers used short-hairpin RNAs to knock down synaptogyrin-3 in the neurons, P301S tau produced a diffuse staining pattern along the axon, suggesting it had largely exited presynaptic terminals. Endogenous mouse tau in wild-type neurons had a similar axonal distribution, which was not affected by knockdown of synaptogyrin-3.

How would mutant tau’s liaison with synaptogyrin-3 affect synaptic function? The researchers transduced the hippocampal mouse neurons with a fluorescently tagged version of the presynaptic vesicle marker synaptophysin, so they could monitor the movement of vesicles in response to stimulation. In neurons from wild-type mice, a 30-Hz zap jacked up vesicle mobility as observed by a drop in fluorescence intensity around the terminals. In neurons from PS19 animals, vesicles remained clustered and immobile, but knocking down their synaptogyrin-3 expression restored the mobility of synaptic vesicles back to normal.

Finally, the researchers measured neurotransmitter release in response to stimulation. Like wild-type neurons, PS19 neurons released an initial burst of synaptic vesicles upon stimulation, but, unlike wild-type, they were unable to sustain it. As stimulation continued, they evoked weaker and weaker excitatory postsynaptic currents from adjacent neurons. Again, knockdown of synaptogyrin-3 rescued this impaired neurotransmitter release.

The researchers drew the same conclusions from similar experiments in fruit flies that express various mutants of tau in motor neurons. Notably, fluorescent recovery after photobleaching (FRAP) experiments in fly motor neurons confirmed that tau’s association with synaptogyrin-3 rendered synaptic vesicles immobile. The researchers concluded that pathological mutants of tau mislocalized to presynaptic terminals, used synaptogyrin-3 to adhere to vesicles, clustered them together, and slowed their release.

“This is an entirely novel mechanism that makes a lot of sense in terms of synaptic dysfunction on the presynaptic side,” commented Karen Gylys of University of California, Los Angeles. “But the hypothesis is a bit difficult to reconcile with some of the features of tau pathology in AD. For example, flow cytometry of total tau labeling in cortical synaptosomes clearly shows abundant tau protein in control as well as AD synapses, suggesting that synaptic tau is not necessarily mislocalized” (Sokolow et al., 2015). She added that tau is phosphorylated, not mutated, in AD.

While McInnes thinks that tau crowding the terminals tips the balance toward synaptic dysfunction, he agreed with Gylys that small amounts of tau may occur normally at the synapse, as detected in postmortem samples from healthy controls.

The researchers suggested that tau-coated synaptic vesicles might propagate the transsynaptic spread of tau between neurons. This would mesh with previous reports that neurons release tau when stimulated (Feb 2014 news). McInnes noted that synaptogyrin-3 also appears on exosomes—tiny vesicles that pinch off of the membrane in close proximity to presynaptic terminals. Mislocalized tau could be transported via those compartments, as well, he predicted. He proposed that small molecules or nanobodies that reduce tau’s association with synaptogyrin-3 might hold therapeutic promise.

Zweckstetter pointed out that tetraspanin vesicle proteins are highly abundant components of synaptic vesicles. “Given the potentially important role of a direct interaction between tau and synaptogyrin-3, it will be important to understand this interaction at the molecular level, in particular in context of the three-dimensional structure of tetraspanin vesicle proteins, which is still enigmatic.” he wrote.—Jessica Shugart

Comments

The study by McInnes et al., together with a previous study from the same group (Zhou et al., 2017), suggests a very interesting mechanism of tau-mediated neurotoxicity. The proposed mechanism is based on a direct interaction of tau with synaptic vesicles, which cross-links synaptic vesicles with filamentous actin and thus affects the mobility of synaptic vesicles at presynaptic compartments. In this new study, a transmembrane protein is identified that mediates the interaction between tau and synaptic vesicles and thus could be an important player in tau-mediated neurotoxicity. The identified transmembrane protein is synaptogyrin-3, a member of the family of tetraspanin vesicle proteins. Tetraspanin vesicle proteins are highly abundant components of synaptic vesicles. Little is known, however, about their function. Given the potentially important role of a direct interaction between tau and synaptogyrin-3, it will be important to understand this interaction at the molecular level, in particular in context of the three-dimensional structure of tetraspanin vesicle proteins, which is still enigmatic.